It has previously been shown that dissolved oxygen in biological fluids can quench the phosphorescence of certain phosphorescent molecules exposed to the fluids, and that it is possible to measure the oxygen concentration by measuring the quenching of phosphorescence. Oxygen quenching may be used, for example, for the non-invasive (apart from the injection of a solution of the phosphorescent agent), quantitative determination of oxygen pressure in the vasculature of tissue in vivo. Commonly invented and assigned U.S. Pat. No. 5,837,865 (Vinogradov et al.) discloses phosphorescent molecules that can be used for imaging of the distribution of dissolved oxygen by imaging the phosphorescence of the molecules when exposed to a suitable source of exciting light.
Oxygen quenching reduces both the intensity and the phosphorescence lifetime or decay time of the phosphorescent light. Commonly invented and assigned U.S. Pat. No. 6,701,168 (Wilson et al.) describes a method of measuring the phosphorescence lifetime by the “phase method” in which a phosphorescent sample is repeatedly excited with a periodic pulsed light source. Each pulse of exciting light causes a pulse of phosphorescence, delayed slightly after the exciting pulse. Thus, the periodic exciting pulse train causes a periodic phosphorescent pulse train at the same frequency, but wherein each pulse is delayed. The delay time, which is a measure of the phosphorescence lifetime, is observed as a phase shift between the two pulse trains.
U.S. Pat. No. 5,127,405 (Alcala et al.) describes a process for determining the intensity/time curve of the phosphorescence in which the sample is excited with a periodic pulsed light source and the detected phosphorescence is measured briefly at intervals slightly greater than the period of the exciting light source. Assuming that all exciting pulses and all phosphorescence pulses are identical, each successive measurement measures the intensity of the phosphorescence at a slightly later time after the exciting pulse, enabling the intensity/time curve to be reconstructed.
Both the methods of the Wilson '168 patent and the method of Alcala rely on combining measurements from a substantial number of successive pulses to form a single image with a time dimension. As a result, these methods can produce results only after a delay. These methods rely on the assumption that the system under observation does not change during the period of observation. Thus, there has, until the present invention, been a need for a method and system for monitoring phosphorescence quenching that can produce accurate images in real time showing a system changing during the period of observation.